Center for Integrated Protein Science (CIPSM) and Zentrum für Pharmaforschung, Department Pharmazie, Ludwig-Maximilians-Universität München, 80539 Munich, Germany.
J Biol Chem. 2012 Aug 3;287(32):26506-12. doi: 10.1074/jbc.M112.357129. Epub 2012 Jun 19.
Activation of hyperpolarization-activated cyclic nucleotide-gated (HCN) channels is facilitated in vivo by direct binding of the second messenger cAMP. This process plays a fundamental role in the fine-tuning of HCN channel activity and is critical for the modulation of cardiac and neuronal rhythmicity. Here, we identify the pyrimidine cyclic nucleotide cCMP as another regulator of HCN channels. We demonstrate that cCMP shifts the activation curves of two members of the HCN channel family, HCN2 and HCN4, to more depolarized voltages. Moreover, cCMP speeds up activation and slows down deactivation kinetics of these channels. The two other members of the HCN channel family, HCN1 and HCN3, are not sensitive to cCMP. The modulatory effect of cCMP is reversible and requires the presence of a functional cyclic nucleotide-binding domain. We determined an EC(50) value of ∼30 μm for cCMP compared with 1 μm for cAMP. Notably, cCMP is a partial agonist of HCN channels, displaying an efficacy of ∼0.6. cCMP increases the frequency of pacemaker potentials from isolated sinoatrial pacemaker cells in the presence of endogenous cAMP concentrations. Electrophysiological recordings indicated that this increase is caused by a depolarizing shift in the activation curve of the native HCN current, which in turn leads to an enhancement of the slope of the diastolic depolarization of sinoatrial node cells. In conclusion, our findings establish cCMP as a gating regulator of HCN channels and indicate that this cyclic nucleotide has to be considered in HCN channel-regulated processes.
超极化激活环核苷酸门控 (HCN) 通道的激活在体内通过第二信使 cAMP 的直接结合来促进。这一过程在 HCN 通道活性的微调中起着基本作用，对于心脏和神经元节律性的调节至关重要。在这里，我们确定嘧啶环核苷酸 cCMP 是 HCN 通道的另一种调节剂。我们证明 cCMP 将 HCN 通道家族的两个成员 HCN2 和 HCN4 的激活曲线移向更去极化的电压。此外，cCMP 加快了这些通道的激活和失活动力学。HCN 通道家族的另外两个成员 HCN1 和 HCN3 对 cCMP 不敏感。cCMP 的调节作用是可逆的，需要有一个功能性的环核苷酸结合域。我们确定 cCMP 的 EC(50)值约为 30 μm，而 cAMP 的 EC(50)值约为 1 μm。值得注意的是，cCMP 是 HCN 通道的部分激动剂，效力约为 0.6。cCMP 在存在内源性 cAMP 浓度的情况下增加了分离的窦房结起搏细胞中起搏电位的频率。电生理记录表明，这种增加是由于激活曲线的去极化移位导致的，而这种去极化移位又导致窦房结细胞的舒张去极化斜率增强。总之，我们的发现确立了 cCMP 作为 HCN 通道的门控调节剂，并表明在 HCN 通道调节的过程中必须考虑这种环核苷酸。
